This laboratory is appropriately titled ?Translational Research,? as we do animal research in normal rodents and in rodents that are genetically altered to mimic the human diseases of the retina. In turn, laboratory findings inform us of opportunities to investigate human disease intervention. We study the cell biology of human and animal retinal tissue using light and electron microscopy, immunohistochemistry, biochemistry, photochemistry, molecular biology and measure retinal function in live animals using the electroretinogram (ERG) and behavior. We focus on 5 main areas.[unreadable] [unreadable] (1) Disease mechanisms and gene therapy for X-linked juvenile retinoschisis (XLRS). XLRS is a leading cause of juvenile macular degeneration in males. As we move towards human gene therapy, our focus is on understanding the molecular interactions, biochemistry and sub-cellular localization of the RS protein and gene regulation. We have localized the protein to particular cell membrane sites that give us clues as to its function using careful anatomical analysis including immunogold electron microscopy and highly sensitive immunohistochemistry with epitope unmasking. We have also discovered molecular interactions which may hold the key to role of RS in the retina. To understand and be able to control RS gene expression, we are isolating and characterizing the human gene promoter region and have identified important regulatory sites. We contine to explore the efficacy of viral gene transfer in restoring normal structure and function in the RS gene knockout mouse at difference ages and have made a detailed study of long term disease progression in this model and long term recovery after treatment. We found that this model mimics some aspects of the disease in humans that have only recently been recognized through more careful and long term clinical observation.[unreadable] [unreadable] (2) Screening for defects in retinal function and morphology in other murine genetic models to explore the possible role of these genes in normal and diseased retina. Mutations in a gene coding for an enzyme involved in the elongation of Very Long chain fatty acids (ELOVL4) causes STGD3 disease in humans, a form of macular degeneration characterized by deterioration in central vision. Combining our studies from both ELOVL4 heterozygous knockout (loss of the gene) mice and knockin (replacement of the gene with a mutated one) mice we found that STGD3 probably does not result from haploinsufficiency, that is, loss of function due to reduced levels of the protein, but from a dominant-negative mechanism where mutated gene product interferes with the normal one. This conclusion is important in guiding further work on the causes of this and other forms of macular degeneration.[unreadable] [unreadable] Usher?s syndrome involves another form of severe retinal degeneration for which no good animal models have yet been devolved. Our investigation of the retinal histology and function in mice with mutations of the Pcdh15 gene, which cause Usher?s syndrome type 1F in humans, did not show retinal degeneration, but did find a consistent reduced ERG response. In addition, a novel splice variant was found which may substitute for the mutated form in mice, but not in humans. This may be the reason Pcdh15 mutations in mice do not cause retinal degeneration. The ERG results indicate that these variants may not completely replace the function of the original gene. Together the findings will lead to work on the functional role of Pcdh15 and possible ways of treating this retinal degeneration.[unreadable] [unreadable] (3) Pharmacological and gene delivery methods of treating retinal degeneration. Besides gene therapy for XLRS, we are investigating other ways of slowing retinal degeneration by broad spectrum types of treatments with growth and neurotrophic factors. We have cloned the gene for two such factors, lens epithelial derived growth factor (LEDGF and heat shock protein 27 (HSP27)), and inserted them into viral vectors for delivery to the retina in rodent models of retinal degeneration. Both agents slowed the retinal degeneration an animal model with inherited disease, the RCS rat. By analyzing the gene expression in these retinas compared to untreated retinas we expect to gain an understanding of the mechanism of disease and protection.[unreadable] [unreadable] (4) Normal and abnormal signaling through retinal neuronal pathways using the electroretinogram and bioactive molecules for modulating retinal responses. We have developed an improved method of blocking specific channels and receptors in the retina to explore their contribution to the ERG and retinal signaling. We injected antibodies to retinal potassium channels to study their effect on the ERG response in the RCS rat, which has an abnormal response from the inner portion of the retina. The antibodies to the channels produced a decline in the response allowing us to deduce that these potassium channels are involved in generating the abnormal response. This approach gives us an opportunity to probe the mechanism of disease states which alter ERG responses, and retinal potassium regulation. [unreadable] [unreadable] (5) Human clinical trials for retinal disease treatment. In 2005 we successfully completed phase I clinical trials for treatment of retinitis pigmentosa (RP) using ciliary neurotrophic factor (CNTF) delivered by encapsulated cell technology (ECT). Treatment involved implantation of ECT devices secreting CNTF into the vitreous chamber of the eye of 10 patients. The study showed no harmful effect of treatment and a proportion of the treated eyes with severe retinal degeneration developed improved visual acuity during the study which remained up to 6 months after the devices were removed. We have now developed protocols for phase II efficacy trials for AMD (below) and RP.[unreadable] [unreadable] Human Protocol 03-EI-0033. X-Linked Juvenile Retinoschisis - Clinical and Molecular Studies. (PI: P.A. Sieving). A genotype-phenotype study of XLRS which results in splitting of the retinal layers. A better understanding of XLRS development might lead to improved treatments through gene transfer.[unreadable] [unreadable] Human Protocol 03-EI-0179. Investigation of the Effect of Dietary Docosahexaenoic Acid (DHA) Supplementation on Macular Function in Subjects with Autosomal Dominant Stargardt-Like and Autosomal Recessive Stargardt Macular Dystrophy. (PI: P.A. Sieving). Evaluate DHA supplementation to improve macular function in patients with Stargardt and Stargardt-like macular dystrophies. DHA fatty acid is essential for normal brain and eye development. [unreadable] [unreadable] Human Protocol 03-EI-0234. A Phase I Study of NT-501-10 and NT-501-6A.02, Implants of Encapsulated Human NTC-210 Cells Releasing Ciliary Neurotrophic Factor (CNTF), in Patients with Retinitis Pigmentosa. (PI: P.A. Sieving). Evaluate safety of CNTF implant in the human eye of retinal degeneration subjects. CNTF protects against retinal degeneration in animal models. Study addresses a major treatment challenge of delivery directly into the human eye. Published: PNAS (2006) Ciliary neurotrophic factor (CNTF) for human retinal degeneration: Phase I trial of CNTF delivered by encapsulated cell intraocular implants.[unreadable] [unreadable] Human Protocol 03-EI-0255. Pilot Study on the Effect of Vitamin A Supplementation on Cone Function in Retinitis Pigmentosa (RP). (PI: P.A. Sieving). A protocol to evaluate whether high dose oral vitamin A will improve retinal function acutely in patients with RP.[unreadable] [unreadable] Human Protocol 06-EI-0071. Phase II Study of Implants of Encapsulated Human NTC-201 Cells Releasing Ciliary Neurotrophic Factor (CNTF), in Participants with Visual Acuity Impairment Associated with Atrophic Macular Degeneration (PI: P.A. Sieving). Evaluate the safety and effectiveness of ciliary neurotrophic factor (CNTF) implants on vision in participants with atrophic macular degeneration. CNTF protects against retinal degeneration in animal models.